Src/ERK but not phospholipase D is involved in keratinocyte growth factor-stimulated secretion of matrix metalloprotease-9 and urokinase-type plasminogen activator in SNU-16 human stomach cancer cell

We investigated the signaling pathway for keratinocyte growth factor (KGF)-induced invasion using human stomach cancer cell line, SNU-16. Alterations in the activities of Src, extracellular signal-regulated kinase (ERK), and phospholipase D (PLD) were measured using [gamma-(32)P] ATP for autophosphorylation of Src, phospho-specific ERK antibody, and [9,10-(3)H] myristic acid, respectively, while herbimycin A, PD98059 and butan-1-ol were used to inhibit their activities. Matrix metalloproteases (MMPs) and urokinase-type plasminogen activator (uPA) were quantified with zymography and Matrigel-coated Transwell was employed to estimate the invasiveness of SNU-16 cells. Src, ERK, and PLD were activated in response to KGF treatment, and inhibition of these enzymes - by their specific inhibitors - decreased KGF-induced invasion in a dose-dependent manner. However, only inhibition of Src and ERK could block KGF-stimulated secretion of uPA and MMP-9. Src, ERK, and PLD are suggested as mediators of KGF-induced invasion in SNU-16. uPA and MMP-9 are considered as downstream targets of Src and ERK whereas PLD is thought to utilize different pathways.